Project 2: Class A pathogens (Anthrax, Plague, Tularemia, Smallpox) resurface as bioterrorism agents or which potent countermeasures must be established urgently. Our basic understanding of the immunopathogenesis of human infectious diseases has always been hampered by the obvious limitation of our inability to directly challenge humans with most pathogens, access to organs that are targeted by the pathogens and the genetic variability within the human population. Therefore, development of new therapeutic antibiotics and vaccines was possible in the context of virus and/or bacteria with high natural incidence in the human population. However, the infectious agents associated with the new biothreats have a very low natural incidence in humans, particularly the inhaled route of these infections. This considerably limits the use of "high risk" populations for testing of therapeutic regimen efficacy. Moreover, this strategy is also becoming limited for a variety of emerging viruses. Therefore, new approaches are required in order to provide the highest level of confidence that the new vaccines and immunomodulators are effective in humans. We now know that vaccines act through DCs and that many pathogens do indeed target DCs though very little information is available about Class A pathogens in that regard. We will thus determine how Class A pathogens alter human DCs in vitro and in vivo upon delivery into the lung. Aim 1 will determine the effect of category A biothreats on human myeloid DC subsets in vitro. Aim 2 will determine the effect of category A pathogens on human plasmacytoid DCs in vitro. Aim 3 will determine the effect of category A pathogens on human DCs in vivo in mice reconstituted with human CD34+HPCs (Humouse). Aim 4 will determine whether Humouse can generate primary and/or secondary immune response to category A pathogens. This project will also establish in vitro and in vivo biosignatures of Category A pathogens and thus complement the database of Category B-C pathogens generated in the Technical Development component.